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Method of preparing 23,24-dihydrocucurbitacin B and use of the same in medicament for treating tumour

A technology of cucurbitacin and dihydrogen, which is applied in the field of medicine, achieves the effects of high-efficiency preparation, inhibition of tumor cell proliferation, and large amount of preparation

Inactive Publication Date: 2008-02-27
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At the same time, 23,24-dihydrocucurbitacin B has the effect on human gastric cancer cell line NUGC-3, human nasopharyngeal carcinoma cell line HONE-1 [4] , human lung adenocarcinoma A549, ovarian cancer SK-OV-3, melanoma SK-MEL-2, central nervous system tumor XF498 and colon cancer HCT15 [6] etc. have certain cytotoxicity, but for its effect on breast cancer cell line Bcap37, breast cancer cell line MCF-7, liver cancer cell line SMMC-7721, colorectal cancer cell line SW620, erythroleukemia cell line K562 and cervical cancer cell line HeLa However, the inhibitory effect has not been reported, nor has the mechanism of its inhibition of tumor growth by inducing tumor cell apoptosis been studied

Method used

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  • Method of preparing 23,24-dihydrocucurbitacin B and use of the same in medicament for treating tumour
  • Method of preparing 23,24-dihydrocucurbitacin B and use of the same in medicament for treating tumour
  • Method of preparing 23,24-dihydrocucurbitacin B and use of the same in medicament for treating tumour

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1: High-purity preparation of 23,24-dihydrocucurbitacin B

[0028] Firstly, the fresh Trichosanthes root (3950g) was lyophilized and then extracted with petroleum ether (2L) and ethyl acetate (3L). The resulting ethyl acetate extract was removed from the solvent at low temperature to obtain an ethyl acetate extract. Then get 2.5g ethyl acetate extract and carry out preliminary separation with three-axis vertical countercurrent chromatography (developed by Zhejiang University Siyuan Natural Medicine and Biotoxin Research Center, column capacity 600ml), and the two-phase solvent system used is sherwood oil, chloroform, Acetonitrile (volume ratio, 6:1:3), then the fractions were combined. Finally, the cytotoxic components in the collected components are prepared by C8 or C18 reversed-phase high-performance liquid chromatography, and the mobile phase is 10%-100% methanol-water gradient elution, the target substance is collected, and the solvent is volatilized to ob...

Embodiment 2

[0029] Example 2: 23,24-Dihydrocucurbitacin B inhibits the proliferation of human tumor cell lines

[0030] experimental method

[0031] (1) Human tumor cell lines and sources: A total of six tumor lines were made, namely breast cancer Bcap37, breast cancer MCF-7, cervical cancer HeLa, liver cancer SMMC-7721, colorectal cancer SW620, and erythroleukemia K562, all from Zhejiang University Cancer Institute.

[0032] (2) 23,24-Dihydrocucurbitacin B was dissolved in DMSO to make 1 mg / ml mother solution, placed in -20°C refrigerator, and diluted with culture solution to final concentrations of 0, 0.6, 1.1, 2.2, 4.5, 8.9, 35.7μM for use, MTT (AMRESCO company product) with phosphate-buffered saline (Phosphate-buffered Saline referred to as PBS) to prepare a 2mg / ml mother solution, placed in a 4 ℃ refrigerator for later use.

[0033](3) Cell culture: all cells are cultivated in RPMI-1640 complete medium, (SIGMA), add 10% newborn bovine serum (Hangzhou Sijiqing Bioengineering Materia...

Embodiment 3

[0039] Example 3: 23,24-Dihydrocucurbitacin B inhibits the apoptosis-related mechanism of breast cancer Bcap37 proliferation

[0040] (1) DNA LADDER experiment: After some cells are treated with drugs, the DNA will be cut by nucleases at the junction of nucleosomes, resulting in DNA fragments of 180-200bp or integer multiples in size, which appear regular in agarose gel electrophoresis This is the most typical feature of apoptosis, and it is also an important sign to distinguish apoptosis from necrosis. The whole experimental process was carried out according to the operation steps of the DNA apoptosis band extraction kit (Biovision). Observe the experimental results under the light.

[0041] The results ( FIG. 4C ) indicated that 3.6 μM 23,24-dihydrocucurbitacin B could significantly induce the apoptosis of breast cancer Bcap37 at 24h, 36h and 48h and produce characteristic DNA ladder-like bands.

[0042] (2) Apoptotic morphology experiment: after breast cancer Bcap37 was co...

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Abstract

The invention discloses a preparing method of 23,24-dihydrocucurbitacin B and usage to treat tumor disease medicine, which comprises the following steps: using adverse current chromatogram; proceeding primary separation for acetic acid ethyl ester extract of trichosanthes anguina root; choosing two phase dissolvent system from light petroleum, chloroform and acetonitrile; merging and collecting the compound with cell toxic property; proceeding against phase high effective liquid phase color spectrum preparation with flowing phase as methanol and water gradient elution; collecting goal matter; volatilizing dissolvent; getting white crystalline powder; getting the product. This invention can be used to prepare the medicine of breast cancer, liver cancer and large intestine cancer, which possesses wide prospect.

Description

technical field [0001] The present invention relates to the technical field of medicine, in particular to a method for extracting active ingredients from plant materials and its use, especially a method for extracting 23,2-dihydrocucurbitacin B from the root of Trichosanthes kirilowli Maxim and its Use in the preparation of antitumor drugs. Background technique [0002] 23,24-dihydrocucurbitacin B (23,24-dihydrocucurbitacin B) is a cucurbitacin substance widely distributed in a variety of plants, and its molecular formula is C 32 h 48 o 8 , molecular weight 560, its chemical structure is as follows: [0003] [0004] People at Wilbrandia ebracteata Cogn. [1] , Cayaponia tayuya [2，3] , Begonia nantornsis [4] , Bryonia Verrucosa [5] , and Trichosanthes Kirilowli [6] The distribution of 23,24-dihydrocucurbitacin B was found in plants such as Trichosanthes L. However, the traditional method for separating and preparing 23,24-dihydrocucurbitacin B such as normal phase...

Claims

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Application Information

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IPC IPC(8): C07J9/00A61K31/575A61K9/00A61P35/00
Inventor 吴世华杨璐刘非燕王国强吴平
Owner ZHEJIANG UNIV
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